Electric fields can be used to create pores in cells without causing permanent damage. This allows for insertion of large molecules into cell cytoplasm. Genes and other molecules such as pharmacological compounds can be incorporated into live cells through a process known as electroporation. The genes or other molecules are mixed with the live cells in a buffer medium. Short pulses of high electric fields are applied to make the cell membranes transiently porous so that the genes or molecules can enter the cells and modify the genome of the cells.
Studies have shown that large size nucleotide sequences (e.g., up to 630 kb) can be introduced into mammalian cells via electroporation (Eanault, et al., Gene (Amsterdam), 144(2):205, 1994; Nucleic Acids Research, 15(3):1311, 1987; Knutson, et al., Anal. Biochem., 164:44, 1987; Gibson, et al., EMBO J., 6(8):2457, 1987; Dower, et al., Genetic Engineering, 12:275, 1990; Mozo, et al., Plant Molecular Biology, 16:917, 1991). However, the efficiency of electroporation, as reflected in the current literature, is usually low (see U.S. Pat. No. 5,019,034, herein incorporated by reference). A typical result is from about 5 to 20 percent transfection depending on conditions, parameters and cell type. Creation of a high efficiency method and apparatus for the of transfer of nucleic acid and the introduction of other preselected molecules into living cells via electroporation is desired.
Genetronics, Inc, San Diego, Calif., has provided an ex vivo flow through electroporation method and chamber in U.S. Pat. Nos. 5,676,646 and 5,545,130, the disclosures of which are incorporated herein by reference.